Identification of the ATP-binding site in the terminase subunit pUL56 of human cytomegalovirus

Scholz, Brigitte; Rechter, Sabine; Drach, John C.; Townsend, Leroy B.; Bogner, Elke

doi:10.1093/nar/gkg229

Cited by 68 publications

(70 citation statements)

References 40 publications

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“…To determine the functional effects of the MLH1 D132H substitution, we expressed the N terminus of MLH1 (amino acids 1-344) containing the ATPase and ATP binding domains with and without this sequence variant in bacteria as a glutathione S-transferase (GST) fusion protein. Consistent with previous studies, GST alone had essentially no ATPase activity, whereas the wild-type MLH1 N terminus fused to GST had measurable activity [8][9][10] . The MLH1 D132H substitution attenuated, but did not eliminate, ATPase activity (Fig.…”

supporting

confidence: 89%

The MLH1 D132H variant is associated with susceptibility to sporadic colorectal cancer

et al. 2004

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Most susceptibility to colorectal cancer (CRC) is not accounted for by known risk factors. Because MLH1, MSH2 and MSH6 mutations underlie high-penetrance CRC susceptibility in hereditary nonpolyposis colon cancer (HNPCC), we hypothesized that attenuated alleles might also underlie susceptibility to sporadic CRC. We looked for gene variants associated with HNPCC in Israeli probands with familial CRC unstratified with respect to the microsatellite instability (MSI) phenotype. Association studies identified a new MLH1 variant (415G→C, resulting in the amino acid substitution D132H) in ∼1.3% of Israeli individuals with CRC self-described as Jewish, Christian and Muslim. MLH1 415C confers clinically significant susceptibility to CRC. In contrast to classic HNPCC, CRCs associated with MLH1 415C usually do not have the MSI defect, which is important for clinical mutation screening. Structural and functional analyses showed that the normal ATPase function of MLH1 is attenuated, but not eliminated, by the MLH1 415G→C mutation. The new MLH1 variant confers a high risk of CRC and identifies a previously unrecognized mechanism in microsatellite-stable tumors. These studies suggest that variants of mismatch repair proteins with attenuated function may account for a higher proportion of susceptibility to sporadic microsatellite-stable CRC than previously assumed.To identify variant alleles in MLH1, MSH2 or MSH6 with high sensitivity and specificity, we designed a new high-density oligonucleotide array (HNPCC Chip) that uses improved bioinformatic

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supporting

confidence: 89%

The MLH1 D132H variant is associated with susceptibility to sporadic colorectal cancer

et al. 2004

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“…Motif 2 in the second cytoplasmic loop is proposed as the glycerol-phosphate binding site based on the glycerol-phosphate K m defects associated with the PlsY[G102A] and PlsY[G103A] mutants. This motif is similar to the ATP binding sites of prototypical kinases that have phosphate binding loops consisting of a glycine-rich sequence followed by a lysine residue (44,45). The glycines are conserved because the presence of side chains would interfere with substrate binding, consistent with the K m defects observed in the PlsY[G102A] and PlsY[G103A] mutants.…”

Section: Glycerol-3-phosphate Acyltranferasesmentioning

confidence: 59%

Thematic Review Series: Glycerolipids. Acyltransferases in bacterial glycerophospholipid synthesis

Zhang

Rock

2008

Journal of Lipid Research

121

122

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Phospholipid biosynthesis is a vital facet of bacterial physiology that begins with the synthesis of the fatty acids by a soluble type II fatty acid synthase. The bacterial glycerol-phosphate acyltransferases utilize the completed fatty acid chains to form the first membrane phospholipid and thus play a critical role in the regulation of membrane biogenesis. The first bacterial acyltransferase described was PlsB, a glycerol-phosphate acyltransferase. PlsB is a key regulatory point that coordinates membrane phospholipid formation with cell growth and macromolecular synthesis. Phosphatidic acid is then produced by PlsC, a 1-acylglycerolphosphate acyltransferase. These two acyltransferases use thioesters of either CoA or acyl carrier protein (ACP) as the acyl donors and have homologs that perform the same reactions in higher organisms. However, the most prevalent glycerol-phosphate acyltransferase in the bacterial world is PlsY, which uses a recently discovered acyl-phosphate fatty acid intermediate as an acyl donor. This unique activated fatty acid is formed from the acyl-ACP end products of the fatty acid biosynthetic pathway by PlsX, an acyl-ACP:phosphate transacylase. Bacterial phospholipid synthesis is a vital facet of bacterial physiology, and the phospholipid head group structures found in the bacterial world come in a truly bewildering variety (1). Phosphatidic acid is a universal intermediate in the biosynthesis of these membrane glycerophospholipids in eubacteria, and this review focuses on the two acyltransferase steps that are common reactions in all glycerophospholipid biosynthesis in bacteria, the glycerol-phosphate and 1-acylglycerol-phosphate (LPA) acyltransferases. These enzymes sit at the interface between the soluble type II fatty acid biosynthetic pathway and the creation of a phospholipid molecule that drives membrane expansion. This pivotal position makes the glycerol-phosphate acyltransferases key regulators of both fatty acid and phospholipid synthesis and has spurred considerable research into the function, selectivity, and regulation of the acyltransferase systems. This review will cover the two acyltransferase systems involved in bacterial glycerophospholipid synthesis, the origin and utilization of acyl donors by these pathways, and their roles in regulating membrane biogenesis. ACYL DONORS IN ACYLTRANSFERASE REACTIONSThe most important acyl donor in bacterial glycerolipid synthesis is acyl-acyl carrier protein (ACP). ACP is a 9 kDa protein that is the acyl group carrier in type II fatty acid synthesis and shuttles the intermediates attached to the sulfhydryl group at the terminus of its 4′-phosphopantetheine prosthetic group between the pathway enzymes (2, 3). These acyl donors are the end products of the bacterial dissociated type II fatty acid synthesis pathway, and in most bacteria, type II fatty acid synthesis is the sole source of fatty acids for membrane phospholipid synthesis. An acyl-ACP intermediate has two possible fates. It can reenter the fatty acid elongation cycle...

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“…The human cytomegalovirus (HCMV) terminase consists of a large and a small subunit, called viral protein pUL56 and pUL89, respectively. The large subunit endows the terminase with ATPase activity and the small subunit is required for cleavage of concatemeric DNA (Bogner, 2002;Hwang & Bogner, 2002;Scheffczik et al, 2002;Scholz et al, 2003). In addition to the terminase, the other critical packaging component is the portal protein.…”

mentioning

confidence: 99%

Assembly of monomeric human cytomegalovirus pUL104 into portal structures

Holzenburg

Dittmer

Bogner

2009

Journal of General Virology

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In order for human cytomegalovirus (HCMV) to replicate, concatemeric DNA has to be cleaved into unit-length genomes and packaged into preformed capsids. For packaging to take place and DNA to be translocated, a channel is required in the capsid. Viral capsid channels are generally formed by portal proteins. Here, we show by cross-linking, native gel electrophoresis of infected cells and gel permeation chromatography that the HCMV portal candidate protein pUL104 can form dimers and higher order multimers. Electron microscopy of purified monomeric pUL104 after 5 min incubation revealed that the protein had assembled into a multimeric form and that this form closely resembles complete portal assembly. This is the first study to show that pUL104 monomers have the ability to form portal complexes without additional viral proteins.The maturation of herpesviruses, as well as doublestranded DNA (dsDNA) bacteriophages, is the first step in the viral life cycle and harbours a number of common features. Starting with the assembly of the viral procapsid, subsequent DNA replication leads to concatenated DNA that is cleaved into unit-length genomes and packaged into the preformed capsids (Adelman et al., 2001;Black, 1989;Bogner, 2002;Catalano, 2000). Packaging is an energyconsumptive mechanism and with bacteriophages it was demonstrated that one molecule of ATP is required to package 2 bp of DNA (Guo et al., 1987;Morita et al., 1993). The viral nanomotor, which is involved in this process, is composed of the terminase and the portal. The human cytomegalovirus (HCMV) terminase consists of a large and a small subunit, called viral protein pUL56 and pUL89, respectively. The large subunit endows the terminase with ATPase activity and the small subunit is required for cleavage of concatemeric DNA (Bogner, 2002;Hwang & Bogner, 2002;Scheffczik et al., 2002;Scholz et al., 2003). In addition to the terminase, the other critical packaging component is the portal protein. With bacteriophages and herpes simplex virus (HSV) type-1, the portal protein forms a homo-multimeric complex that serves as a channel for translocation of the DNA into the capsid (Valpuesta & Carrascosa, 1994;Newcomb et al., 2001Newcomb et al., , 2005. In addition to this, the portal plays a fundamental role in the assembly of capsids. Even though the portal proteins known so far share no sequence homology, the architecture of the portals is strikingly similar. All portal proteins occupy one vertex location in the capsid and are thought to be dodecamers upon integration (Guasch et al., 2002; Lurz et al., 2001;Trus et al., 2004;Valpuesta & Carrascosa, 1994); they are typically composed of a crown, a stalk and a wing domain. The crown and the stalk delineate a narrow central channel between which the wider wing domain is located (Agirrezabala et al., 2005;Poliakov et al., 2007).With HCMV, previous studies have suggested that the open reading frame UL104 encodes the portal protein . In vivo, pUL104 directly interacts with the large terminase subunit pUL56, which was sho...

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Identification of the ATP-binding site in the terminase subunit pUL56 of human cytomegalovirus

Cited by 68 publications

References 40 publications

The MLH1 D132H variant is associated with susceptibility to sporadic colorectal cancer

The MLH1 D132H variant is associated with susceptibility to sporadic colorectal cancer

Thematic Review Series: Glycerolipids. Acyltransferases in bacterial glycerophospholipid synthesis

Assembly of monomeric human cytomegalovirus pUL104 into portal structures

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